1887

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Volume 70, Issue 7

Adaptation of and assays for diversity profiling by Illumina sequencing to identify and No Access

Abstract

Bartonellosis is an emerging zoonotic disease caused by bacteria of the genus . Mixed infections are a well-documented phenomenon in mammals and their ectoparasites. The accurate identification of species in single and mixed infections is valuable, as different species have varying impacts on infected hosts.

Current diagnostic methods are inadequate at identifying the species present in mixed infections.

The aim of this study was to adopt a Next Generation Sequencing (NGS) approach using Illumina sequencing technology to identify species and demonstrate that this approach can resolve mixed infections.

We used Illumina PCR amplicon NGS to target the and genes of in fleas collected from cats, dogs and a hedgehog in Israel. We included artificially mixed samples to demonstrate the ability for NGS to resolve mixed infections and we compared NGS to traditional Sanger sequencing.

In total, we identified 74 , two two and three fleas. Real-time PCR of a subset of 48 fleas revealed that twelve were positive for , all of which were cat fleas. Sanger sequencing of the and genes confirmed the presence of , and . Illumina NGS of and amplicons further confirmed the species identity in all 12 flea samples and unambiguously resolved the artificially mixed samples.

The adaptation and multiplexing of existing PCR assays for diversity profiling via NGS is a feasible approach that is superior to traditional Sanger sequencing for speciation and resolving mixed infections. The adaptation of other PCR primers for Illumina NGS will be useful in future studies where mixed bacterial infections may be present.

  • Received:
  • Accepted:
  • Published Online:
Keyword(s): Bartonella , Ctenocephalides , Diagnostics , Identification and Next Generation Sequencing
© 2021 The Authors
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2021-07-23
2024-04-25
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Adaptation of gltA and ssrA assays for diversity profiling by Illumina sequencing to identify Bartonella henselae, B. clarridgeiae and B. koehlerae
J. Med. Microbiol. 70, 001400 (2021); https://doi.org/10.1099/jmm.0.001400
/content/journal/jmm/10.1099/jmm.0.001400
/content/journal/jmm/10.1099/jmm.0.001400
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